The interaction phytochemicals from Broccoli with gold salt results in dual reduction and surface capping to produce well-defined stable and biocompatible gold nanoparticles. The core sizes of B-AuNPs are in the range of 10–20 nm as measured from TEM techniques which was also corroborated by CPS technique. Hydrodynamic diameter of B-AuNPs is 100±5 nm, suggesting that broccoli phytochemicals are capped on gold nanoparticles. The negative zeta potential of -29.0 mV for B-AuNPs indicates that the particles repel each other and there is no tendency for the particles to aggregate. The order of cytotoxic activity of B-AuNPs in terms of IC-50 value was as follows: MDA-MB-231 (160 µg/ml)> PC-3 (150 µg/ml)> U266 (125 µg/ml)> SkBr3 (80 µg/ml)> T47D (22 µg/ml). The cellular localization of B-AuNPs against breast and prostate cancer cell lines was evaluated using dark field optical microscopy and transmission electron microscopic image analysis (Image 1). B-AuNPs showed synergistic effects and excellent internalization of prostate (PC-3) and breast (MDA-MB-231 and T47D) cancer cells at concentrations of 25 and 50 µg/ml respectively. This unique synergistic cocktail effect of B-AuNPs may provide new opportunities for generating biocompatible AuNPs for applications in molecular imaging and therapy.